The present invention, in some embodiments thereof, relates to pharmaceuticals and more particularly, but not exclusively, to novel cochleate-based systems and uses thereof as drug delivery vehicles in the treatment of medical conditions such as, for example, conditions associated with antibiotic-resistant pathogenic microorganisms.
Antibiotic resistance represents a worldwide health problem where treatment failure of an ever increasing number of pathogens is intimately associated with severe outcomes such as increased mortality and morbidity. This grave state of events is largely due to a multitude of biochemical and genetic strategies that bacteria have developed to neutralize the threats imposed by antibiotics. Resistance acquirement by bacteria can be divided into four main aspects: i) direct inactivation by hydrolysis, acylation or oxidation; ii) target modification that reduce sensitivity to antibiotics (e.g. ribosome structure; iii) target bypass, i.e., bacteria become refractory to specific antibiotics by bypassing their inactivation mechanism; iv) efflux pumps, which reduce intracellular drug concentration due to their active export out of the cell. While efflux pumps may induce relatively low level resistance to many classes of antibiotics, especially macrolides, tetracyclines, and fluoroquinolones, which inhibit biosynthesis and therefore must accumulate inside bacteria, they contribute significantly to multidrug resistance (MDR).
Host derived cationic antimicrobial peptides (AMPs) and their synthetic mimics are widely regarded as a potential source of future therapeutic agents against a broad range of pathogens. Various AMPs have shown an ability to act synergistically with conventional antibiotics such as β-lactams, macrolides, tetracycline, ciprofloxacin and rifampin, thereby sensitizing antibiotic-resistant bacteria. Conversely, antibiotics may sensitize AMP-resistant bacteria although, in all cases, the molecular basis for these phenomena was not addressed experimentally. While peptide-based antimicrobials represent a class of promising agents in fighting bacterial resistance to antibiotics, difficult challenges need to be overcome towards their eventual use in therapeutics, including the need to improve bioavailability, toxicity and production costs. Another concern for therapeutic uses of AMPs pertains to the emergence of extreme resistance phenomena to the host defense system. Bacteria may sense AMPs via a variety of two-component sensor/regulator systems (TCSs, e.g., PhoPQ or PmrAB), that regulate specific gene expression leading to greater stability of the outer membrane and adapting bacteria for survival. As some AMPs were found to activate one or more TCSs, concern stems from general use of host defense peptides which may provoke the evolution of resistance that will compromise our natural defenses against infection. Hence, synthetic AMP-mimics that retain antibacterial activity but lack the ability to activate PhoQ or its orthologs would represent preferable therapeutics.
A wide range of strategies were put forward in the attempt to alleviate one or more of these shortcomings through chemical mimics that reproduce the AMPs most critical biophysical characteristics in unnatural, sequence-specific oligomers.
U.S. Pat. No. 7,504,381, WO 2006/035431, WO 2008/132738, WO 2009/090648 and U.S. Patent Application Nos. 20070032428 and 20100120671, by one of the present inventors, which are incorporated herein by reference as if fully set forth herein, teach a novel class of peptidomimetic antimicrobial and/or anticancerous polymers. These antimicrobial and/or anticancerous polymers, also referred to as oligo-acyl-lysyl (OAK) polymers, are composed of hydrophobic moieties and amino acids, and maintain three key attributes of AMPs: a flexible structure, an amphiphatic character and a net positive charge. As presented in these patent applications, these antimicrobial polymers are composed of positively charged amino acid residues, such as lysine, and non-amino acid hydrophobic moieties, such as ω-amino-fatty acid residues, as well as fatty acid residues, which not only achieve the desired amphiphatic trait and resolve the production and maintenance issues limiting the use of polypeptides as drugs, but also alleviate the severe limitations restricting the administration of polypeptides as drugs.
The aforementioned OAK polymers have been shown to exhibit high and synergistic antimicrobial and/or anticancerous activity, low resistance induction, re-sensitization of antibiotic-resistant pathogens, non-hemolyticity, resistibility to plasma proteases and high affinity to microbial membranes. Hence, the OAK polymer approach appears to offer advantages owing to its simplicity of design which so far generated OAK sequences with selective antimicrobial and antitumor properties both in test tubes and in animal models of disease, while exhibiting a certain potential for addressing problems related to MDR phenomena.
Oral delivery is the most suitable way of administering drugs for most non-hospitalized, non-acute care patients. Drug delivery systems that allow oral delivery improve patient compliance and facilitate treatment outside the hospital, which has a significant impact on healthcare economics. Many drug delivery platforms have emerged and are present in either a preclinical stage or in an advanced clinical trial with the intent of trying to demonstrate efficient oral absorption. In particular, cochleate technology was shown to be effective in the therapeutic oral delivery of the hydrophobic antifungal agent amphotericin B.
Cochleates are roll-like microstructures that consist of a series of lipid bilayers, which are formed as a result of the condensation of small unilamellar negatively charged liposomes. In the presence of calcium, the small phosphatidylserine (PS) liposomes fuse and form large sheets which have hydrophobic surfaces and, in order to minimize their interactions with water, tend to roll-up into the turbinated cylindrical lipid bilayers, or cochleate.
Cochleates were discovered in 1975, and have been used in the 80s and 90s to transport antigens and peptides for vaccine delivery. It was demonstrated that by using a binary phase system, such as two non-miscible hydrogels, cochleates can be formed that display a small mean particle of less than 500 nm. These cochleates were highly suitable for the encapsulation of hydrophobic drugs, such as amphotericin B.
Freeze-fracture electron microscopy reveals a typical cochleate cylinder characterized by the elongated shape and by the tight packed bilayers. Because cochleates contain both hydrophobic and hydrophilic surfaces, they are suitable to encapsulate both hydrophobic drugs like amphotericin B and clofazimine and amphiphathic drugs like doxorubicin. The loading efficacy of the cochleates depends upon the physical chemistry of the drug to encapsulate, whereas the particle size of the drug-cochleate complex depends on the process used to encapsulate. The main components of currently known cochleates are phosphatidylserine (PS) and calcium, two natural compounds. Phosphatidylserine is a constituent of the brain and is sold in health stores as a nutrient supplement.
WO 1996/025942 and WO 1997/030725 disclose cochleates comprising a biologically relevant molecule component, a negatively charged lipid component, and a divalent cation component, wherein the biologically relevant molecule can be a polynucleotide or a polypeptide.
U.S. Pat. No. 6,592,894 discloses a process for producing a small-sized, lipid-based cochleates which are derived from liposomes that are suspended in an aqueous two-phase polymer solution, and treated with positively charged molecules such as Ca2+ or Zn2+, and which may contain biologically relevant molecules.
U.S. Patent Application No. 20040092727 teaches cochleates wherein the agents bridging the lipid bilayer are organic multi-valent cations such as 2,3,5,6-tetraminopyrimidine sulfate.
Syed, U M, et al. [Syed, U M, et al., Int J Pharm, 2008, 363, (1-2):118-125] disclose cochleates which are able to microencapsulate water-soluble cationic drugs or peptides into its inter-lipid bi-layer space. These cochleates were formed through interaction between negatively charged lipids and drugs or peptides, such as poly-L-Lysine, acting as the inter-bi-layer bridges in addition to the presence of Ca2+.